1. Field of the Invention
The present invention relates to Coryneform bacteria containing plasmids made by recombinant DNA techniques carrying genetic information useful for the fermentative production of lysine.
2. Description of the Prior Art
L-lysine, which is used as a feedstuff, is normally produced industrially by a fermentation process in which L-lysine producing mutants of the genus Brevibacterium or Corynebacterium are used. Various L-lysine-producing mutants, produced by the artificial mutation of wild strains of microorganisms of the genera Brevibacterium and Corynebacterium, are known. Examples of such artificial mutants are mutants resistant to S-(2-aminoethyl)-cysteine (hereinafter identified as AEC), mutants requiring L-amino acids such as L-homoserine for their growth (Japanese Published Examined Patent Application Nos. 28078/1973, 6499/1981), mutants resistant to AEC and further requiring an amino acid such as L-leucine, L-homoserine, L-proline, L-serine, L-arginine, L-alanine or L-valine, (U.S. Pat. Nos. 3,708,395 and 3,825,472), L-lysine-producing mutants resistant to DL-.alpha.-amino-.epsilon.-caprolactam, .alpha.-amino-lauryllactam, aspartic acid-analogues, sulfa drugs, quinoids, N-lauroyl leucine, or to inhibitors of oxalacetate, decarboxylase or respiratory system enzymes (Japanese Published Unexamined Patent Application Nos. 53588/1975, 31093/1975, 102498/1977, 9394/1978, 86089/1978, 9783/1980, 9759/1980, 32995/1981, 39778/1981, Japanese Published Examined Patent Application Nos. 43591/1978, 1833/1978), L-lysine-producing mutants requiring inositol or acetic acid (Japanese Published Unexamined Patent Application Nos. 9784/1980, 8692/1981), and L-lysine-producing mutants sensitive to fluoropyruvic acid or to temperatures higher than 34.degree. C. (Japanese Published Unexamined Patent Application Nos. 9783/1980, 86090/1978). Copending, coassigned U.S. patent application Ser. No. 333,455, now U.S. Pat. No. 4,411,977 filed Dec. 22, 1981, discloses a mutant of Brevibacterium or Corynebacterium which is resistant to ethylene glycol and which produces L-lysine.
The prior art has described Escherichia coli strains transformed with a recombinant plasmid DNA, which strains have increased productivity of amino acids (see, for example, U.S. Pat. No. 4,278,765).
Although the presence of plasmids in strains of Brevibacterium and Corynebacterium having detectable phenotypic markers has not been known for a long time (but see, e.g., Published European Patent Application No. 003391), recent work has demonstrated the feasibility of preparing Coryneform bacteria harboring plasmids controlling the production of threonine or isoleucine see the commonly assigned, Ser. No. 376,396 now U.S. Pat. No. 4,452,890 copending at the U.S. Patent and Trademark Office, filed May 10, 1982 (thr), and commonly assigned copending Ser. No. 392,145, now U.S. Pat. No. 4,442,208 filed June 25, 1982 at the U.S. Patent and Trademark Office (ile)). In addition, commonly asigned copending U.S. patent application Ser. No. 386,980, now U.S. Pat. No. 4,514,502 filed on June 10, 1982 at the U.S. Patent and Trademark Office, describes composite plasmids capable of propagating in Coryneform glutamic acid producing bacteria. (All of these patent applications are herein fully incorporated by reference.)
A brief schematic representation of the lysine, isoleucine and threonine biosynthetic pathway is as follows: ##STR1##
The first branch of the pathway leading to threonine and isoleucine can be found at the junction of aspartate semialdehyde. The enzyme leading to these two amino acids is homoserine dehydrogenase (hereinafter "HDase"), while the enzyme leading to lysine is dihydrodipicolinate synthetase, ("DHDP synthetase").
It should also be noted here that, recently, Escherichia coli strains were described which carry hybrid plasmids containing several genes of the lysine biosynthetic pathway. An overproducer of lysine (TOC R 21) was transformed, and the synthesis of lysine was studied in different strains (see, for example, (LeReverend at al, European Journal of Applied Microbiology and Biotechnology, 15: 227-231 (1982), as well as published French Patent Application No. 2511032 (Application No. 81/15385) published Feb. 11, 1983. It appears from these publications that only plasmids containing the dapA gene (encoding DHDP synthetase) lead to an increase in lysine production; this reaction is the limiting biosynthetic step in lysine overproducers (having mutations altering the aspartokinase reaction). The authors suggest that such a method of gene amplification could be used to improve strains which overproduce metabolites. There is no suggestion in these publications, however, to expand this work to Cornyeform bacteria. Commonly assigned copending U.S. application, Ser. No. 504,471, now U.S. Pat. No. 4,601,983 filed on June 15, 1983, discloses recombinant plasmids carrying genetic information to control isoleucine and threonine biosynthetic pathways, present in Cornyeform bacteria.
A need still exists for improved and efficient methods for the fermentative production of L-lysine.